During the maturation of ovarian follicles, FSH is known to induce the synthesis of a variety of proteins in addition to its well-known effects on steroidogenesis. The endocrinology of follicular development has been studied by investigators in several laboratories and granulosa cells have been shown to be the principle direct target of the gonadotropin. Further, a variety of agents including estrogens, androgens, LHRH, adenosine, and others are known to influence the response of granulosa cells to FSH, particularly in culture. Beyond the observation that cyclic AMP can mimic most of the actions of FSH on stimulation of steroidogenesis and expression of several proteins (i.e., aromatase, LH receptors, and plasminogen activator, among others), little is known about the mechanism used by FSH and cyclic AMP in this process. Progress in understanding hormonal effects on gene expression in other systems has depended on the use of cloned probes to specific genes which are induced in response to hormonal stimulation. These probes are not yet available for the study of FSH induced gene expression in granulosa cells. Our first specific aim will be to prepare several DNA probes which can be used to study FSH induced granulosa cell gene expression. Once the probes are made and appropriately characterized, our second specific aim will be to use them to study the mechanism by which FSH and cyclic AMP initiate expression of these genes. Estradiol is well known to act synergistically with FSH in inducing several proteins including one (i.e., aromatase) that enhances its own formation. Therefore, we will also use the probes to investigate mechanisms by which FSH and estradiol act synergistically. Further, we will study the influence of a variety of other agents known to influence the responsiveness of estradiol to FSH in culture including LHRH, calcium ionophores, and phorbol esters. Our third specific aim, an important longer range goal of this application, is to isolate and characterize the genes for aromatase and LH receptors. These are FSH-induced proteins with key endocrine roles in the process of follicular maturation. Their low abundance suggests that we will need to use antibodies as aids in preparation of cDNA probes. Whereas antibodies to the aromatase are available, we plan to prepare monoclonal antibodies against the receptor.